Cancer Metastasis: Building a framework

1. Cancer Metastasis: Building a framework Gaorav P. Gupta and Joan Massague Gabriella F. de Paz and Susana S. Hak Fall 2011, 20.309

2. An Evolutionary Metaphor • Evolution = environmental stresses select for organisms with advantageous survival traits • Metastasis = cellular and micro-environmental stresses select for tumor cells with advantageous survival traits • Both situations require genetic heterogeneity: evolution  meiosis metastasis  genomic instability

3. Take-home: Successful metastasis requires TCs to face and overcome a wide array of biological defenses put forth by the body.

5. Intrinsically Advantageous Molecular Mechanisms • altered cellular adhesions • loss of E-cadherin-mediated adhesions • deregulated cell motility • gain-of-function mutations in Rho family GTPases • resistance to extracellular death signals • ectopic overexpression of anti-apoptotic effectors (i.e. BCL2) • tumor-initiating capacity • overexpression of transcriptional repressor Bmi-1

6. Beyond the Basement Membrane • disruption of the basement membrane and ECM • deregulation of ECM proteases • co-option of stromal cells • gene expression signature of fibroblast activation in vitro indicates which cancers are more likely to metastasize • recruitment of immune response cells • tumor-associated macrophages secrete vasoactive factors and GFs

8. Intravasation Metastatic tumor cells  cancer-associated vasculature  lymph nodes  hematogenous circulation Advantageous molecular mechanisms: • Twist: TF that promotes EMT + rate of hematogenousintravasation • Motility: motile tumor cells can freely move down natural chemoattractive gradient

9. Stresses Encountered in Transit • physical damage from hemodynamic shear forces • immune-mediated killing • anoikis(debatably) Circumvention Mechanisms: • co-opt blood platelets • overexpression of BDNF-receptor trkB

10. Extravasation Timing and method of escape into vasculature and target tissue varies: • tumor cells grow in intravascular space until lesion bursts through surrounding vasculature • tumor cells release ezrinas an anchoring mechanism (osteosarcoma) • tumor cells release signals that permeabilize vasculature (ex. VEGF)

11. Colonization Two theories on target tissue selection: • path-based theory: • target tissue of CTC is determined by the TCs circulatory pattern and immediate need to adhere • surface interaction (honing) theory: • target tissue of CTC is determined by molecular interaction of ectopically expressed ligand-receptor pairings • research indicates latter theory holds more merit Dormancy period: • some TCs lose proliferative ability upon entering target tissue • limiting factors vary among tumors • known as minimal residual disease (MRD)

13. Site-Specific Colonization: Bone • Preservation of bone homeostasis • Osteoclasts Osteoblasts • Two types of cancer cells metastasize to bone • Breast cancer: hyperactivation of osteoclasts • Prostate cancer: simulation of osteoblasts • Positive feedback loop • “The vicious cycle of osteolytic bone metastasis” • Bone-metastatic signature manifested in breast cancer

14. Site-Specific Colonization: Lungs • Common with metastatic diseases • Cardiac output circulates through lung-capillary network • Initiated through small pulmonary arterioles • Burst through or breach endothelial junctions and basement membrane • Growth factors in breast cancers implicated in lung metastasis • Same breast-cancer cell line associated to bone metastasis • Gene-expression signature enriched with mediators

15. Site-Specific Colonization: Brain • Less frequent and poor prognosis • Colonize brain parenchyma or thrive along the leptomeninges • Blood brain barrier (BBB)protects the central nervous system • Compromised in metastasis • Lack of reliable experimental models

16. Site-Specific Colonization: Liver • Densely vascularized tissue • Vessels highly porous for circulating cells and nutrients • Rate-limited step for metastasis • Invasion in hepatic parenchyma • Avoidance of cell death from immune cells • Hepsin protease promote liver metastasis of prostate cancer

17. Seeding and Reseeding • Genes promoting metastasis coexpressed within subset of primary tumors • Selectable growth advantage • Metastatic cells may travel back to point of origin • Intrinsic colonizing function  constantly reseeding primary tumor • Link with large tumor size, rapid growth rate, and metastatic behavior

18. Impact of the Cell of Origin • Certain cell lineages expresses small molecules that bias the metastatic efficiency to different target organs • Development history of cell can cause activation of specific metastasis-promoting mechaisms • Impact of developmental predisposition  transformation occurs at different stages within same lineage • Organism level predisposition  cell is mutated but is phenotypically silent

19. Overview • Metastasis, a somatic evolution from cancerous cells • Heterogeneity allows selection for advantageous traits  overcome environmental defenses • Primary tumors thrive indicative of stromal-cell path • By understanding metastasis and with new advancing technologies can lead to new avenues for clinical therapy

20. Supplemental Slides

21. Co-option of Stromal Cells • co-option of stromal cells by tumor cells have been implicated in increasing a cell's metastatic ability: • disrupting TFGß signaling between fibroblasts can induce carcinomas in certain organs in mice: shows that interfering with tumor-suppressing crosstalk in stromal cells can potentially lead to metastatic advantages in non-stromal tumor cells • looking at gene expression signature of fibroblast activation in vitro gave indication of which cancers were more likely to metastasize • breast cancer associated fibroblasts make chemokine CXCL12, which augments the proliferation and migratory activity of tumor cells + facilitates angiogenesis

22. Recruitment of Immune Response Cells • inflammatory cells from the immune system synthesize prostaglandins (pro-metastasis) • tumor-associated macrophages secrete copious amounts of vasoactive factors (VEGF, IL-8, etc) that potently induce angiogenesis and proteases that enhance their biological activity • macrophages also release GFs that facilitate survival, proliferation, and invasion during cancer progression (evidence: mice with defects in macrophage production seldom produce metastasizing carcinomas from aggressive mammary tumors)

23. Homing Examples • Homing of disseminated tumor cells to a secondary organ may be a result of rapid lodging into capillaries (i.e. cell not wanting to be un-adhered for too long) but recent studies propose that it's more specific than that: • could be an adhesive interaction between cell-surface receptors expressed on malignant cells (integrins) and their cognate ligands expressed on various target sites (adhesive proteins) in target site for metastasis (ex: a3ß1----> laminin-5 on exposed basement membrane in lung metastasis) • could also be interaction between ectopically expressed chemokines and their receptors (ex: in breast cancer cells, CXCR4 directed metastasis to CXCL12-rich tissues like lungs).

24. Confirmations of Extravasation Examples • cytoskeletal anchoring protein ezrin aids extravasation in osteosarcoma cells (inhibiting ezrin's expressions in these cells incited higher rates of cancer cell death prior to successful escape into lung parenchyma) • certain signals emanating from metastatic cells induce vascular permeability + make it easier for tumor cells to invade (VEGF has been shown to do this - activates Src family kinases in endothelial cell junctions and causes disruptions - proven by experiment that saw Src knockout mice protected from VEGF-secreting cancer cell metastasis)

25. Generating a Viable Niche • mobilization of hematopoietic progenitors from the bone marrow via circulation and into target sites for metastatic colonization • regulated/released in response to hormonal factors emitted by the primary TCs • hematopoietic cells found to express VEGFR1, CD133, CD34, and c-kit in target tissue • “Targeted inhibition of VEGFR1-expressing progenitors using neutralizing antibodies suggested that this preconditioning was necessary for metastatic progression.” • “A subcutaneously inoculated lung carcinoma that induced these bone marrow-derived progenitors to congregate only in the lungs also metastasized only to that site, whereas a melanoma that recruited these progenitors to multiple organ sites exhibited a widespread metastatic tropism.”